The presence and concentration of target biological analytes, such as, nucleic acids or proteins, in a sample fluid containing one or more other molecules can be determined by using the complex binding of these target biological compounds with probes. For example, a set of specific probes, each of which is chosen in order to interact specifically (i.e. hybridize) with one particular target analyte, can be immobilized at specific locations of a biosensor solid substrate. The target analytes are usually labeled with a detectable label molecule (e.g., a colored particle, a fluorophore, or an electrochemically active moiety). By contacting the solid substrate with the sample fluid, the target analytes are fixed at the locations corresponding to their specific probes. The detection of the target analytes in the sample fluid is then operated via the localization of the signals produced by the detectable molecules bound to the target analytes.
Because hybridization is a temperature-dependent phenomenon, temperature control provides significant advantages in this technology, e.g., for nucleic acid analyses. Moreover, nucleic acid analytes typically require amplification prior to detection, and most amplification techniques involve maintaining the fluid sample at one or more temperature(s) for a predetermined amount of time in order to achieve reliable amplification. Thus, accurate temperature control is also important for analyte processing steps preceding detection.
The ability to measure temperature changes of liquids in very small volumes within a microfluidic environment presents a number of technical challenges. Usually, the measurement requires interactions with the liquid by measuring devices which either provide poor contact due to distance and surface area available for measurement or result in heat transfer between the solution and the measuring device that may interfere with accurate measurement. Thus, there is a need for improving, in a cost-effective manner, the temperature control in a microfluidic environment, especially up to the level where substantially a homogeneous temperature within a few tens of degrees Celsius can be achieved. There is a need for a precise and reliable method and device to monitor the temperature, and its distribution, directly at the level of the biosensor substrate. There is also a need for a method of making such improved devices, such that the method is relatively easy to perform and does not significantly increase the cost of the device.
Accordingly, it is an object of the present invention to provide improved methods that permit accurate monitoring and/or control of temperature changes in a microfluidic environment. An advantage of the present invention is that the temperature can be monitored and/or controlled at any location within a microfluidic device where a preparation step, an amplification step or a detection step is performed. A further object of the invention is to provide improved devices for monitoring and/or controlling temperature in a microfluidic environment that can be employed to practice the methods disclosed and claimed herein.